Animal experiments have demonstrated that cytomegalovirus (CMV)-vectored vaccines are unique in that they: a) induce and maintain high frequencies of extralymphoid T cell responses (so called effector memory T cells); b) super-infect CMV-positive hosts; and c) maintain immunogenicity even when rendered deficient in host-to-host spread. Furthermore, experiments in animal models have shown that vaccine vectors derived from animal CMVs induce a protective immune response against infectious diseases and cancer (US 20080199493; US 20100142823; US 20130136768; and US 20140141038; all of which are incorporated by reference herein). Particularly striking is the finding that a rhesus CMV (RhCMV)-vectored simian immunodeficiency virus (SIV)-vaccine was able to not only prevent AIDS in non-human primates, but ultimately cure these animals from SIV (Hansen S G et al., Nature 502, 100-104 (2013); incorporated by reference herein).
It is important to use an attenuated strain in the development of a cytomegalovirus vaccine because an unattenuated strain could spread from host to host and potentially be pathologic at least in immunocompromised individuals. Previously, attenuated human CMV (HCMV) strains have failed to a) establish latent infection (Plotkin S A and Huang E S, J Infect Dis 152, 395-397 (1985); incorporated by reference herein); b) induce long-lasting immunity (Jacobson M A et al., J Clin Virol 35, 332-337 (2006); incorporated by reference herein); c) reinfect the significant proportion of the population that has been previously naturally infected with CMV (Heineman T C et al., J Infect Dis 193, 1350-1360 (2006); incorporated by reference herein); or d) produce persistent infections (WO2013/036465; incorporated by reference herein.) Furthermore, clinical strains of HCMV genomes are highly unstable in vitro when grown in fibroblasts, resulting in fibroblast adaptations such as deletion of UL131A.
The impact of such adaptations to tissue culture for the ability to perform vector functions in vivo is mostly unknown. In addition to the need for attenuations to be stable in vitro and in vivo, it is important that these vectors can be manufactured with reproducible results. The most stable attenuation strategy is gene deletion. However, this generally requires the generation of complementing cell lines which is difficult to achieve for primary cells used to grow cytomegalovirus.